This invention relates to a composition and method for inhibiting the activity of terminal deoxyribonucleotidyl transferase (TdT).
At the present time, there are four clasess of DNA mammolian polymerases which function to promote the production of DNA cells. Three of the DNA polymerases, known as alpha, beta or gamma DNA polymerase, require a template strand of DNA and a primer strand of DNA in order to effect the addition of substrates on the primer strand by addition of monophosphate residues at the 3-prime-OH end of the primer strand according to the Watson-Crick base pairing rule as dictated by the template strand. These DNA polymerases are known as replicative DNA polymerases. TdT polymerase differs from replicative DNA polymerases in that it requires only a single stranded DNA initiator molecule upon which to initiate synthesis. The TdT catalyzes the polymerization of deoxyribonucleotide on a 3-prime-OH end of oligo or polydeoxyribonucleotide initiators in the absence of the template.
Under normal conditions, TdT is present only in thymus and bone marrow with the highest concentrations being found in the thymocytes. From this observation, it has been postulated that TdT plays a role in immunodifferentiation, at least in the T-cell series. It has also been reported by McCaffrey et al in Proc. Acad. Sci. USA, vol. 70, No. 2, pp. 521-525, February 1973, that TdT is present in lymphoblastic leukemia cells. Furthermore, it has been found that TdT is not present in myeloblastic leukemia cells or lymphosarcoma-cell leukemia. While all patients having acute lymphoblastic leukemia do not test positively for TdT, the vast majority, about 95% of such patients, test positively for TdT. Accordingly, conventional biochemical techniques or fluorescent antibody labeling techniques for the TdT marker can be utilized to provide at least a preliminary test to determine the presence or absence of acute lymphoblastic leukemia cells in patients.
It has been proposed to utilize selective inhibitors of replicative DNA polymerases in order to inhibit replication of gram-positive bacteria. 6-(arylhydrazino) and 6-(arylamino) uracils have been found to be selective inhibitors of DNA replication by inhibiting bacterial DNA polymerase III. It is believed that the mechanism involves the specific pairing of substituents of the uracil moiety with template cytosine and binding of the 6-aryl group and its substituents to the polymerase, thereby sequestering the polymerase in a relatively stable protein:drug complex. It has been found that only a narrow spectrum of enzymes--the type III polymerases of gram-positive bacteria is susceptible to these uracil inhibitors. This is because the polymerase III possesses, at a critical location near the active site, a unique aryl site which strongly binds a 6-aryl moiety. It has been shown that the compound 6-(p-n-butylanilino) uracil is a specific inhibitor of DNA polymerase alpha of HeLa leukemia cells and, in vivo, is a selective inhibitor of HeLa cell division and DNA synthesis. No other mammalian cell DNA polymerase is inhibited by this compound.
It would be desirable to provide inhibitors of TdT in order to inhibit the growth of and/or cause the destruction of lymphoblastic leukemia cells in vivo or in vitro.